Optically-active amino acids in high purity are becoming increasingly important as intermediates for the preparation of pharmaceuticals, foods and agrochemicals. Hoppe et al., Chemie in Unserer Zeit, 17, 41-53 (1983); and Kleeman et al., in Ullmann's Encyclopedia of Industrial Chemistry, Vol. A2, pages 57-97.
For example, D-phenylglycine and D-p-hydroxyphenylglycine are side chain precursors for the semisynthetic penicillins, ampicillin and amoxycillin, respectively. See Schmidt-Kastner et al., in Biotechnology, Vol. 6a, edited by H. J. Rehm and G. Reed, Verlag Chemie, pages 388-419 (1984). In addition, the most important building block for the low-calorie sweetener, aspartame, is L-phenylalanine. Mazur et al., J. Am. Chem. Soc., 91, 2684 (1969). Moreover, D-phenylalanine possesses analgesic properties and might one day supplant aspirin [Nutrition News, Vol. VI, no. 7 (1983)]; and D-valine is an intermediate for the pyrethroid insecticide, Fluvalinate [Farm. Chem. Handbook, 72nd ed., Meister Publ. Co. (1986)].
Examples of essential amino acids which are important in human nutrition include L-isoleucine, L-leucine, L-lysine, L-methionine, L-phenylalanine, L-threonine, L-tryptophan and L-valine. The uses of these essential amino acids have been described in several review articles. Izumi et al., Angew. Chem. Int. Ed. Eng., 17, 176 (1978); and Drauz et al., ibid., 21, 584 (1982).
For many applications, it is necessary to have an optically active amino acid as the starting material which has an optical purity of at least about 65 percent and preferably at least about 90 percent. Such a purity level appears to be a requirement particularly in the case of optically active acyl amino acids.
There are essentially four processes that are used for the production of amino acids. These comprise: a) extraction; b) chemical synthesis; c) fermentation; and d) enzymatic catalysis.
In the extraction process, the natural L-amino acids are isolated from protein-containing animal and vegetable products. However, because of the high production costs, only the amino acids, L-cystine, L-tyrosine and L-proline can be economically produced using this method. Kleeman et al., Ullmann's Encyclopedia of Industrial Chemistry, Vol. A2, page 67.
In principle, all amino acids can be prepared by chemical asymmetric syntheses. However, the only process that has provided for large-scale production is the production of L-dopa via hydrogenation in the presence of an optically-active rhodium complex. Knowles et al., J. Am. Chem. Soc., 99, 5946 (1977). The more common chemical methods are optical resolution by direct crystallization of enantiomeric mixtures, Collet et al., Chem. Rev., 80, 215 (1980), and fractional crystallization of diastereomeric salt pairs as disclosed in U.S. Pat. No. 4,224,239 to Nippon Kayaku. However, all of these processes are expensive and are not universally applicable.
The fermentation process is used commercially only for the biosynthesis of natural L-amino acids. The fermentation equipment used in the process is expensive. And the lower yields of other amino acids using this process make it economically prohibitive.
The fourth process of amino acid production involves enzymatic catalysis. The general principles of the catalysis are well known in the art and include use of the following enzymes to provide the L-amino acid as one of the reaction products: (1) an L-specific esterase; (2) an L-specific acylase; (3) a nitrile hydratase followed by an amidase; and (4) an aminopeptidase.
However, there are several disadvantages associated with use of an enzymatic catalysis process. Since it is a kinetic resolution procedure, the desired enantiomer can be obtained in only 50 percent theoretical yield. Moreover, tedious separation of the acid from the ester or amide is required. The undesired enantiomer is isolated and racemized again for recycling.
Most enzymatic catalysis processes are designed for the synthesis of a particular amino acid. For example, L-aspartic acid is prepared industrially via stereospecific amination using aspartase from E. coli: L-aspartate in turn can be transformed into L-alanine with an L-aspartate-.beta.-decarboxylase from Pseudomonas dacunhae, Chibata et al., J. Appl. Microbiol.. 13. 638 (1965).
For another example, a combination of chemical and microbiological methods is used for the synthesis of L-lysine. Thus, DL-.alpha.-amino-.epsilon.-caprolactam is obtained from cyclohexane in three steps. Hydrolysis of the L-isomer by cells of Candida humicola containing L-specific .alpha.-amino-.epsilon.-caprolactamase produces L-lysine. The D-.alpha.-amino-.epsilon.-caprolactam is racemized by a racemase from Alkaligenes faecalis, T. Fukesmura, Agric. Biol. Chem., 41, 1321 and 1327 (1977). This process is used by Toray industries for commercial scale L-lysine production.
For an additional example, a biochemical method for the production of L-serine from DL-oxazolidine-4-carboxylic acid is known. This procedure involves the hydrolysis of the L-oxazolidine into L-serine by Pseudomonas testosteroni. The D-oxazolidine is racemized to DL-oxazolidine by a racemase from Bacillus subtilis, Yokozeki et al., Agric. Biol. Chem., 51, 963 (1987).
A Similar enzymatic process was developed for the commercial production of L-cysteine from DL-2-aminothiazoline-4-carboxylic acid (DL-ATC). In this case, both the L-ATC hydrolase and ATC racemase reside in the same microorganism, Pseudomonas thiazolinophilum, Sano et al., Agric. Biol. Chem., 43, 2373 (1979).
A more general biocatalytic asymmetric process is based on the stereospecific action of microbial hydantoinases. In this process, the nonreactive hydantoin antipode is racemized under the reaction conditions or via the action of a racemase. Apparently, this methodology is commercialized only for the production of D-phenylglycine and D-p-hydroxyphenylglycine. The disadvantage of this process is that the hydantoinases are substrate-specific which requires tedious screening to find hydantoinases for the synthesis of each amino acid in the enantiopure form.
A specific enzymatic catalysis process which has received considerable attention over the past several years is the use of a lipase to stereoselectively hydrolyze esters. Lipases are well known and many of these are available commercially. The properties of lipases, in particular their biochemical actions, are described by H. L. Brockman in Lipases edited by B. Borgstrom and H. L. Brockman, Elsevier, Amsterdam, p. 3 (1984). Lipases are used commercially for the transesterification of fats and are incorporated in laundry detergents for removal of oily contaminants.
One example of a catalyzed process using a lipase is disclosed in WO 9015146. Racemic mixtures of esters of 2-substituted acids, other than 2-halopropionic acids, are stereoselectively hydrolyzed in the presence of a lipase of Candida rugosa in an organic solvent. Preferably, the process is carried out in a reducing agent to yield high optical purity.
Another specific enzymatic catalysis process using a lipase is disclosed in Japanese Patent No. 2215391. The lipase of Candida cylindracea was used to hydrolyze N-acyl-amino acid esters to produce glycine. The reaction was carried out at room temperature and the yields were about 70 percent.
The 5(4H)-oxazolones, commonly known as azlactones, and as exemplified by Formula (1) herein, are useful intermediates in the synthesis of .alpha.-amino acids and peptides. They can undergo ring opening with various nucleophiles such as water, alcohols, amines, thiols, amino acid esters, and the like to produce amino acid derivatives.
In principle, if the unreactive azlactone is racemized in situ under the reaction conditions, quantitative transformations of a racemic azlactone by a suitable biocatalyst (enzyme) is possible. The behavior of azlactones towards proteolytic enzymes and carboxyesterase has been examined. However, the reactions were carried out in dilute mixtures because the reaction rate is slow. Moreover, the optical purity of the product is not sufficiently high for industrial use, de Jersey et al., Biochemistry, 8, 1967 (1980); ibid., 9, 1761 (1970); Daffe et al., J. Am. Chem. Soc., 102, 3601 (1980).
In fact, in a very recent publication, Bevinakatti et al., J. Chem. Soc. Chem. Commun., 1091 (1990), it is stated that, "Because azlactones undergo spontaneous hydrolysis in the presence of water, previous attempts (referring to de Jersey et al. and Daffe et al.) to cleave them enantioselectively using enzymes or cyclodextrins as catalysts have met with little success." In an attempt to overcome this problem of spontaneous hydrolysis of azlactones in aqueous media, Bevinakatti et al. carried out their reactions in anhydrous organic solvents. They attempted the enzyme-catalyzed enantioselective ring-opening of 2-phenyl-4-methyl-oxazolin-5-one using butan-1-ol as the nucleophile in diisopropyl ether (DIPE) as solvent.
Bevinakatti et al. found that the lipases of Candida cylindracea (CCL) and porcine pancreatic lipase (PPL) quantitatively transformed the azlactone, 2-phenyl-4-methyl-oxazolin-5-one, into R-butyl-N-benzoylalanine: but the enantiomeric excess (ee) was found to be only 10 percent and 3 percent, respectively. On the other hand the lipase of Mucor miehei converted the same azlactone into S-butyl-N-benzoylalanine with an optical purity of 57 percent ee at 45 percent conversion but reduced to 34 percent ee at 100 percent conversion. These results clearly demonstrate that the Bevinakatti et al. process is unsuitable for producing optically-active acyl amino acids having the foregoing required high optically-active purity levels.
The art needs a new and useful process for producing optically-active acyl amino acids from oxazolone precursors.